The objective is the mechanism by which the B-lactam antibiotics interact with the penicillin-binding proteins (PBPs) and penicillin sensitive enzymes (PSEs) of the bacterial peptidoglycan crosslinking system (PgCS) and inhibit wall synthesis. On the basis of the previous two years of NIH-supported studies, the program includes : (a) the functioning, structure and geometry of the B-lactam active center (presumably a regulatory center) and te L-R-D-Ala-D-Ala active center (i.e., the catalytic one) in the PSEs. Our investigations on the enzyme active centers of model PSEs isolated to protein homogeneity from the culture filtrates of various Actinomycetes will be expanded. Affinity labelling, amino acid sequencing and X-ray crystallographic techniques will be used. The studies will be extended to several highly purified membrane-bound PBP's/PSEs : the streptomyces 20-24,000 dalton DD-transpeptidase, the streptococcus faecalis 43,000 dalton DD-carboxypeptidase and the 40,000 dalton DD-carboxypeptidase of the L-form of Proteus mirabilis. These three selected PBPs/PSEs are likely to be important components of the PgCS in the corresponding bacteria. And (b) the identification, genetic analysis and physiological functions of the various cell-bound PBPs/PSEs (up to 8 in S. faecalis) which together with those alluded to above, are members of the PgCs in the three selected organisms. The program has significance for both clinical (control of bacterial disease; intrinsic resistance) and basic bacteriology.